Staphylococcus aureus (SA) is a major threat to public health with no vaccines available for human use. Currently a large number of SA clinical isolates are methicillin resistant (MRSA) making the development of staphylococcal vaccines a pressing public health need. The pathogenicity of SA is dependent on a plethora of virulence factors including cell surface proteins and polysaccharides as well as toxins that target the immune cells or cause tissue destruction enabling the pathogen to disseminate and seed in distant organs. Several prior attempts to develop vaccines for S. aureus have ignored the importance of key toxins. The myriad of cytolytic and immune modulating toxins cripples the ability of the innate immune response to control the infection leading to invasive disease. Prior failed efforts for development of SA vaccines using surface proteins or polysaccharides (ClfA, SdrG, IsdB, CP5, and CP8) were focused on achieving sterile immunity (total prevention of infection) through an opsonophagocytic mechanism, an approach successfully applied to other major pathogens such as S. pneumoniae. However, the results of the clinical trials with these vaccines (Nabi, Meck, Biosynexus) question the notion that opsonophagocytic antibodies can mediate sterile immunity against S. aureus. Inability to induce effective opsonic antibody response may relate to expression of protein A by S. aureus. This proposal offers an alternative approach by devising a strategy to induce broadly neutralizing antibodies to key S. aureus toxins with the goal of preventing the complications of invasive disease (clinical protection- such as prevention of sepsis) rather than sterile immunity. The proposal is based on a set of structurally designed vaccine candidates for three classes of staphylococcal toxins, i.e. single component alpha hemolysin (Hla), bicomponent leukocidins, as well as superantigens. A further foundation of the proposed study is a prospective clinical study demonstrating a strong inverse correlation between pre-existing antibody titers to the selected toxins and the probability of sepsis in immunocompetent adults with S. aureus bacteremia. The current proposal is aimed at demonstration of efficacy of a multivalent formulation in animal models of S. aureus infection leading to a preclinical multivalent vaccine candidate. The proposal will further explore the mechanism and correlates of the conferred immunity informed by a systematic analysis of the neutralizing epitopes represented in convalescent patients. The proposal is designed in four Specific Aims: In Aim 1 we will develop a bi- or trivalent pore-forming toxoid vaccine formulation with broad neutralizing activity towards Hla and the family of leukocidins and efficacy in mice and rabbits. In Aim 2, a trivalent vaccine formulation will be developed using three superantigen toxoids and efficacy will be demonstrated in a novel humanized mouse model expressing HLA-DR4. One component of this vaccine (STEBVax) is already in Phase I clinical trial under an NIAID supported program (VTEU). In Aim 3 a final pentavalent formulation will be developed including pore-forming and superantigen toxoids and tested in several infection models in HLA- DR4 mice as well as rabbits. In Aim 4, using a large cohort of convalescent and septic patients with S. aureus bacteremia, we seek to determine the relationship between protection from sepsis and presence of antibodies to specific epitopes in Hla and leukocidins. In summary this project completes over a decade of work supported by US government and is expected to result in a novel vaccine for S. aureus as well as potential correlates of immunity to guide clinical development. The proposal brings together three highly skilled teams: Integrated Biotherapeutics with extensive expertise in toxoid vaccines; Diep's lab (UCSF) a pioneer in rabbit models of S. aureus infections, and Sidhu's lab (U Toronto) a leading laboratory in phage display human antibody libraries.